1. Field of the Invention
The present invention is broadly concerned with improved immunoassay processes wherein use is made of a site-deactivating medium for reducing or substantially eliminating non-specific interactions between a sample being tested and the surfaces of the reaction vessel. More particularly, it is concerned with an improved method of this type wherein the vessel wall surfaces in contact with the sample and antigen-antibody components during the reaction are coated and covalently bonded with a medium such as a total biological fluid or extract for site-deactivation purposes.
2. Description of the Prior Art
A number of immunoassay techniques have been proposed in the past for determining, either qualitatively or quantitatively, the presence of a component of an antigen-antibody reaction in a given sample. For example, immunodiffusion and immuno-electrophoresis, complement fixation, passive hemagglutination and radio-immunoassay procedures have been developed. Furthermore, U.S. Pat. No. 3,654,090 describes an immuno-chemical assay process wherein use is of a component of an antigen-antibody reaction in insolubilized form, whereas the other component is covalently linked to an enzyme. The insolubilized and free enzyme-labeled components are added to a sample to be determined, and the antigen-antibody reaction is allowed to proceed to completion. If the sample contains one of the components of the antigen-antibody reaction, this component competes with the corresponding added component for reaction with the other component of the reaction. When the antigen-antibody reaction is complete, the insoluiblized and free fractions are separated, and the activity of the labeled component is determined by an appropriate measure of one of the separated fractions.
Solid-state techniques analogous to that outlined above are in widespread use. In such processes, one of the components of an antigen-antibody reaction is adsorbed onto an insoluble carrier or surface in a reaction vessel; for example, the component can simply be adsorbed directly to the wall surfaces of a synthetic resin vessel. A reaction mixture fraction is then added to the vessel which would include the sample being determined and the other component of the antigen-antibody reaction which has been labeled. Such labeling can be accomplished by a number of means, for example using a radioisotope or covalently linking an enzyme to the free component. The antigen-antibody reaction is then allowed to process, wherein one of the following possibilities can occur: (1) If the sample being determined is free of the labeled component, then all of the latter will react with the insolubilized fraction, and the remaining reaction mixture fraction will be free of antigen; or (2) If the sample contains a quantity of the component corresponding to the labeled component, a competition results between the labeled component and that in the sample. In this case, because of such competition, an amount of the labeled component will remain in the reaction mixture fraction. In either event, the remainder of the reaction mixture fraction and the insolubilized fractions are separated, and the label activity is measured on one of these fractions. Determination of activity of course depends on the nature of the label; in the case of a radioisotope, an isotope counter is employed, whereas if an enzyme label is employed, enzyme activity may be determined colorimetrically.
While the above described solid-state assay procedures are known, a persistent problem which has detracted from the usefulness thereof involves non-specific interaction which can occur between components of the sample and the reaction vessel wall surfaces and/or other adsorption surfaces present during the antigen-antibody reaction. As can be appreciated, such non-specific interaction can materially detract from the accuracy of the solid-state technique, particularly if quantitative determinations are desired.
To give but one example of this problem, many insurance companies today require a urine sample from applicants for their policies, and such samples are often checked for the presence of thiazides therein. If such thiazides are present, a good indication is given as to whether the applicant is taking certain types of medications. In any event, use of the above-described solid-state technique in connection with thiazide determinations presents significant problems relating to non-specific interactions, which is enhanced because of varying quantities of urea present in the urine samples. In fact, such interactions can be so significant as to lead to totally erroneous qualitative results, i.e., a urine sample free of thiazide tests as a positive, or a thiazide-containing sample tests as a negative.
There is therefore a decided need in the art for an improved immunochemical assay process which overcomes the problems associated with non-specific interactions, particularly in the case of solid-state immunoassay techniques.